患者, 男性, 56岁, 因"发热半个月, 腹胀7 d"于2019年5月入住吉林大学第一医院血液科。否认皮肤瘙痒, 咳嗽、咳痰, 腹痛、腹泻等胃肠道症状。既往糖尿病病史8年, 血糖控制可;有土霉素过敏史;吸烟史20余年(已戒20年), 饮酒史30年;近期无旅游史, 无不洁饮食, 未食用生冷海鲜食物。查体:ECOG评分3分, 一般状态差, 无贫血貌, 皮肤及巩膜黄染, 全身浅表淋巴结未触及, 胸骨压痛阴性, 双肺底呼吸音弱, 心脏查体无明显异常, 腹部膨隆, 肝脾肋缘下未触及, 移动性浊音阳性, 双下肢无水肿。血常规:WBC 66.6×109/L, 嗜酸性粒细胞22.25%(14.82×109/L), HGB 158 g/L, PLT 20×109/L;肝功能:谷丙转氨酶39 U/L, 谷草转氨酶48 U/L, 总胆红素73 μmol/L, 直接胆红素39 μmol/L, 白蛋白26 g/L;呼吸道病毒、巨细胞病毒(CMV)、EB病毒(EBV)、降钙素原(PCT)等感染相关检验阴性, 自身免疫系统疾病筛查阴性。浅表淋巴结超声可见颈部、腋下及腹股沟多发淋巴结肿大(最大1.3 cm×0.6 cm, 皮髓结构尚清)。CT示"双侧胸腔及腹腔中等量积液, 肝脏脾脏不大, 腹主动脉旁、腹腔内见多发淋巴结影(0.4～1.4 cm)"。骨髓象:有核细胞增生明显活跃, 粒系增生明显活跃, 原始粒细胞比例增高(0.175), 嗜酸细胞比例明显增高(0.325), 其中幼稚嗜酸性粒细胞占0.065, 粒细胞颗粒明显增多;红系增生活跃;淋巴细胞比例减低, 形态正常。骨髓活检病理:骨髓有核细胞增生极度活跃, 粒红比例增高, 幼稚细胞易见, 嗜酸细胞多见, 网状纤维染色MF-2级灶性。骨髓免疫分型:异常髓系原始细胞占有核细胞4.52%, 主要表达CD34str、CD33、HLA-DR、CD123、CD7、CD11bdim、CD25, 嗜酸性粒细胞比例明显增高(占28.1%)。髓系/淋系肿瘤融合基因筛查:BCR-ABL1等32种融合基因阴性。二代基因测序未检出RUNX1、DNMT3A、TET2等111种髓系/淋系肿瘤相关基因突变。荧光原位杂交(FISH)检测:FGFR1基因重排占94.5%(图1), PDGFRα、PDGFRβ及JAK2基因重排阴性。染色体核型46,XY,-7,t(8;13)(p11.2;q12),+der(13)t(8;13)(p11.2;q12)(?)[18]/46,XY, t(8;13)(p11.2;q12)[2]。RT-PCR及直接测序证实ZMYM2-FGFR1融合基因阳性。临床诊断:伴嗜酸性粒细胞增多和FGFR1重排的髓系肿瘤。

急性髓系白血病(AML)是一组高度异质性疾病,现已 发现约95%的AML患者具有一种以上基因突变,某些基因 突变为患者个体化治疗及靶向治疗提供基础,并且对AML 的预后判断具有重要参考价值。 DNA甲基化修饰是参与 基因表达调控和染色质结构重塑过程重要的表观遗传学机 制,构成 AML 发病的第III类致病基因。DNA 甲基转移酶 3A(DNA methyltransferase 3A, DNMT3A)是一种与表观遗 传修饰相关的重要蛋白酶,和 DNMT3B 共同承担 DNA 从 头甲基化的修饰工作。目前对DNMT3A突变的预后意义仍 存在一定争议,大部分研究者认为DNMT3A突变与AML患 者的不良预后相关,但也有研究提示DNMT3A突变的不 良预后价值可能仅限于特定类型AML患者群体中,仅少 数研究报道DNMT3A突变在整体及特定亚组AML患者中 均不能作为独立预后标志。现回顾性分析我中心初治正 常核型(normal karyotype, NK)AML患者的临床资料,以探 究DNMT3A突变及其伴随突变在NK-AML患者中的预后 价值。

Objective To investigate the current status and real performance of the detection of RUNX1-RUNX1T1 fusion transcript levels and WT1 transcript levels in China through interlaboratory comparison.Methods Peking University People's Hospital (PKUPH) prepared the samples for comparison.That is,the fresh RUNX1-RUNX1T1 positive (+) bone morrow nucleated cells were serially diluted with RUNX1-RUNX1T1 negative (-) nucleated cells from different patients.Totally 23 sets with 14 different samples per set were prepared.TRIzol reagent was added in each tube and thoroughly mixed with cells for homogenization.Each laboratory simultaneously tested RUNX1-RUNX1T1 and WT1 transcript levels of one set of samples by real-time quantitative PCR method.All transcript levels were reported as the percentage of RUNX1-RUNX1T1 or WT1 transcript copies/ABL copies.Spearman correlation coefficient between the reported transcript levels of each participated laboratory and those of PKUPH was calculated.Results ①RUNX1-RUNX 1T 1 comparison:9 samples were (+) and 5 were (-),the false negative and positive rates of the 20 participated laboratories were 0 (0/180) and 5％ (5/100),respectively.The reported transcript levels of all 9 positive samples were different among laboratories.The median reported transcript levels of 9 positive samples were from 0.060％ to 176.7 ％,which covered 3.5-log.The ratios of each sample's highest to the lowest reported transcript levels were from 5.5 to 12.3 (one result which obviously deviated from other laboratories' results was not included),85 ％ (17/20) of the laboratories had correlation coefficient ≥0.98.②WT1 comparison:The median reported transcript levels of all 14 samples were from 0.17％ to 67.6％,which covered 2.6-log.The ratios of each sample's highest to the lowest reported transcript levels were from 5.3-13.7,62％ (13/21) of the laboratories had correlation coefficient ≥0.98.③ The relative relationship of the reported RUNX1-RUNX1T1 transcript levels between the participants and PKUPH was not always consistent with that of WT1 transcript levels.Both RUNX1-RUNX1T1 and WT1 transcript levels from 2 and 7 laboratories were individually lower than and higher than those of PKUPH,whereas for the rest 11 laboratories,one transcript level was higher than and the other was lower than that of PKUPH.Conclusion The reported RUNX1-RUNX1T1 and WT1 transcript levels were different among laboratories for the same sample.Most of the participated laboratories reported highly consistent result with that of PKUPH.The relationship between laboratories of the different transcript levels may not be the same.

Objective: To establish primary immune thrombocytopenia (ITP) animal model induced by anti-platelet membrane glycoprotein GPⅠbα antibodies AN51 and R300. Methods: Twenty guinea pigs (6-8 week) were divided into 4 groups. Five guinea pigs in each group were intravenously injected with different doses of AN51 (0.05, 0.1, 0.2 μg/g) and 0.2 μg/g IgG as control. The whole blood was collected from inner angular venous plexus. Platelets number was determined by an automated cell counter and Swiss-Jim method. Then, the similar protocol was used to establish ITP nude mice model by intraperitoneal injection of different concentrations of anti-platelet GPⅠbα antibody R300, respectively. Results: ①Five minutes after intravenous injection of AN51 at 0.05, 0.1 and 0.2 μg/g, the platelet counts of guinea pigs reduced about 0-5%, 50%-60% and 70%-80% compared to the control group, respectively. The difference was statistically significant (P<0.01) . ②Six hours after intraperitoneal injection of R300 at 0.05, 0.1, 0.2 μg/g, the platelet counts of nude mice decreased about 20%-30%, 60%-70% and 80%-90% compared to the control group, respectively. The difference was statistically significant (P<0.01) . The nude mice, injected 0.2 μg/g R300 once a day for 2 weeks, showed typical ITP clinical manifestations including large number of petechiaes or ecchymoses on limbs, head and abdomen. Conclusion: AN51 at 0.2 μg/g and R300 at 0.2 μg/g could establish stable ITP model in guinea pigs and nude mice respectively.

Retinoic acid (RA), an activated metabolite of vitamin A (all-trans-retinol), controls multiple pathways essential for vertebrate embryonic development and perinatal growth and function. Rdh1 is the most efficient enzyme known (Vm/Km) that catalyzes dehydrogenation of all-trans-retinol, contributes to a reconstituted path of atRA biosynthesis in intact cells, and belongs to a family of enzymes (short-chain dehydrogenases/reductases, SDR) capable of recognizing the physiological form of vitamin A, retinol bound with cellular retinol binding-protein. To further assess the function of Rdh1 in RA homeostasis, the murine gene was disrupted by homologous recombination. Null mice fed a retinol-enriched diet were born in Mendalian ratio, and remained healthy and fertile. They did not present with any of the morphological abnormalities of vitamin A deficiency. The Rdh1 null mouse, however, had a 2-fold greater all-trans-retinol in liver than wild-type. Interestingly, the mRNA of Cyp26A1, a major catalyst of RA degradation, was down-regulated 3-fold in livers of Rdh1 null mice fed a diet with standard or marginal amounts of vitamin A. Thus, the vitamin A normal phenotype was achieved through compensation by decreased expression of the major atRA degrading enzyme in the liver. Consistent with this function for CYP26A1, mRNA expression levels of CYP26A1 and Rdh1 showed a strong inverse relationship in the whole embryo from e7.5 to e18.5 and in the liver from e12.5 to P2M. This is the first genetic evidence for a function of a specific retinol dehydrogenase in a phyisological pathway of atRA generation. This work was supported by NIH grant DK36870.

Protein phosphatase 2A (PP2A) is one kind of serine/ threonine protein phosphatase regulating mainly cell growth and division. It comprises three subunits, A, B, and C. The B-subunit is involved in enzyme activity and substrate specificity. The B-subunit family is of great diversity and is divided into three classes, the B1, B2, and B3 subfamilies. Until now, two members of the B1 subfamily, B1alpha and B1beta, have been identified in human. In this report, the third member of the sub-family, B1gamma, was identified, and its cDNA was isolated from a human brain cDNA library. This novel cDNA is 4,120 bp in length and contains an open reading frame (nt 55-1,398) encoding 447 amino acid residues. The putative protein shares 81 and 85% identity with B1alpha (PPP2R2A) and B1beta (PPP2R2B), respectively, and was named PPP2R2C for its high level of homology to the other two isoforms. One remarkable characteristic of this novel gene is that it is highly expressed in brain with a 4.7-kb transcript while it is nearly undetectable in other tissues. In addition, the PPP2R2C gene was localized to human chromosome 4p16 between markers D4S2925 and D4S3007 with 5.45 cR (LOD > 14) and 2.63 cR (LOD > 15) RH distance, respectively, by radiation hybrid panel mapping.

Using the conservative nucleotide sequences encoding the catalytic domain of the beta-1, 4-GalT genes in human, bovine, mouse, chick and snail as probes to search the NCBI GenBank EST database, several ESTs with high homology were obtained. Primers were designed in the flanking sequence of EST contig. Using the PCR product amplified in human placenta cDNA library as probe to perform "walking" hybridization with human placenta cDNA library, a cDNA fragment with the length of 1,907 bp was cloned. It contained an open reading frame (ORF) with the length of 1,179 bp, which encodes 393 amino acid residues. The deduced amino acid sequence of this gene shares 43.8% identity to the human beta-1, 4-GalTI and 60.9% in the catalytic domain especially. The expression mapping showed that it was expressed in human most tissues with a single 2.4 kb transcript, but the relative expression level of the transcript are vary. While this gene was mapped on chromosome 1 using the cDNA hybridization with human/rodent hybrid cell line DNA Southern blot panel.

The Dhm1 gene is the mouse homologue of the dhp1(+) gene of Schizosaccharomyces pombe, which is involved in homologous recombination and RNA metabolism, such as RNA synthesis and RNA trafficking, in S. pombe. Complementation analysis showed the Dhm1 gene on a multicopy plasmid can rescue the temperature-sensitivity mutation of dhp1(ts) and the lethality of the dhp1 null mutation. This finding suggests that Dhm1 has a function in mouse similar to that of dhp1(+). The human homologue of this gene, XRN2, has been identified. A 3.6-kb transcript of XRN2 was detected in 16 tissues examined and was more abundant in testis. By radiation hybrid panel mapping, the XRN2 gene was localized to chromosome 20p11.1-p11.2 between markers D20S180 and D20S871.

By low stringency PCR amplification of genomic DNA using the primers designed based on the conservation of zinc finger motif, we got 8 gradient eletrophoretic bands. After recovery of the second and third bands, the DNA fragments in them were cloned and sequenced. Compared to the GenBank database, among these 60 segments containing zinc finger motif, 23 segments were novel zinc finger genes' genomic segments. Then the human brain tissue cDNA library was screened, using these segments as probes, and 44 positive clones were obtained. Rescreening 28 of them, we got 20 rescreened clones. All of them were sequenced and sent to the GenBank DNA database for sequence analysis, the results showed that 16 were novel C2H2 type zinc finger protein cDNA segments. The cDNA segments encoding the novel C2H2 type zinc finger proteins provide the basic materials for cloning of full length cDNA of valuable novel zinc finger protein genes.

A pair of degenerate primers were designed according to the DNA sequences of the C(2)H(2) zinc finger genes conservative domain and then homologious PCR was performed with the human genomic total DNA as template. The zinc finger fragments thus obtained were used as probes to screen cDNA libraries of human fetal kidney, muscle and marrow and 22 C(2)H(2) zinc finger gene fragments were selected. Among these fragments, 17 were confirmed to be novel zinc finger gene fragments by literature searches in the National Center Biotechnology Information (NCBI) database. Expression patterns of the clones K3-4 and K5-12 selected from kidney cDNA library were analyzed. The results showed that the expression level of K3-4 in kidney is obviously higher than in other tissues and K5-12 is expressed at different levels in 8 tissues.

The strategy of isolating the band-specific expression fragments from the probe pool of human chromosome generated by microdissection was reported in present paper. A chromosome 14 q 24.3 band-specific single copy DNA library was constructed based on this probe pool. Using this pool DNA as probe to hybridize the human bone marrow cell cDNA library, 68 primary positive clones were selected from 5 x 10(5) cDNA clones. Of them 32 clones were got in second-round screening and designed as cFD 14-1-32. Finally, 24 bandspecific expression fragments were identified from these 32 positive clones by analysing the results of DNA hybridization. Those band-specific clones can hybridize to both 14 q 24.3 DNA and human genomic DNA, but have no hybridization signal with 17 q 11-12 DNA. Partial sequences of 13 fragments of them were sequenced and were identified as novel cDNA sequences as well as have some homology with known genes in NCBI database. Analysis of expression spectrum of cFD 14-1 suggested that the cDNA fragments thus obtained can be used to isolate the genes not yet be cloned in 14 q 24.3 region.